We report here on the anti-CMV activity of ten new artemisinin-derived dimers, one deoxy dimer and one new artemisinin monomer, and the identification of the most potent anti-CMV dimer. The enhanced anti-CMV activity of dimers as compared to monomers was not a result of increased cytotoxcity but rather a specific anti-CMV activity. The two most highly potent dimers were effective in inhibition of the laboratory-adapted Towne strain as well as a GCV-resistant strain. They were also shown to have the strongest inhibitory effect on the growth of cancer cell lines.
All dimers exhibited potent CMV inhibition at 1 µM. However, only two compounds (832-4 and 838) could be selected as highly potent anti-CMV agents based on their high CMV inhibition at 0.1 µM and their low cytotoxicity. At this concentration, the other dimers had either no inhibition or at most 40% inhibition of luciferase expression, while GCV had no inhibition at all.
Artemisinin derivatives were initially developed as anti-malarial drugs, but later were also found to have additional pharmacological activities such as growth inhibition of cancer cells and inhibition of CMV replication 
. Whether or not these new activities are related to each other and indicate a shared mechanism, the endoperoxide bridge appears to be critical in all pharmacological activities of artemisinin derivatives.
As a generalized observation, the dimers were significantly more potent than the monomers in CMV inhibition. Using the EC50
dimer 838 was 165-fold more potent than artesunate in anti-CMV activity and its SI was 134-fold higher compared to artesunate. In support of this finding, the model of dose response slope parameter which was recently used to characterize the activities of anti-HIV compounds 
revealed that dimers had a much higher slope than monomers. Generally, a higher slope correlates with a more vigorous anti-viral activity. This could result either from very high affinity of drug binding to a single ligand or from cooperative effect of multiple ligands with similar affinity.
Several anti-cancer agents (such as kinase inhibitors) have been shown to inhibit CMV replication, but in general, the concentrations required to achieve CMV inhibition were toxic to cells and prohibited their use as anti-viral candidates 
. Interestingly, Sorafenib, a multi-kinase inhibitor, appears to inhibit CMV replication without associated cellular toxicity 
. The dimers reported here prevented growth of cancer cell-lines and were potent inhibitors of CMV replication without apparent cellular toxicities. Therefore, they may provide a new class of anti-CMV agents, with a distinct mechanism of action. Similar to the anti-CMV activity, dimers were also more effective than monomers in inhibition of cancer cells. Future studies may reveal whether the dual anti-CMV and anti-cancer activities of dimers result from a shared mechanism of action. We and others have shown that inhibition of CMV replication with artemisinin derivatives appears early during the replication cycle and involves a mechanism which is different from the DNA polymerase inhibitors 
. The anti-cancer activities of artemisinins have been postulated to involve cell cycle arrest, apoptosis, and/or angiogenesis 
, processes that are also affected by specific CMV genes, mostly immediate early genes. Future studies will address the mechanism of CMV inhibition by artemisinin-derived dimers and their potential overlap with oncogenic signaling events.
In conclusion, we have shown that: 1). Dimers are highly active against CMV replication, and dimer diphenyl phosphate 838 is the most potent in CMV inhibition. 2). The dose response slope parameter of dimers is significantly higher than that of artemisinin monomers and GCV, a possible indication that dimers are a distinct class of anti-CMV compounds. 3) Dimers are more potent inhibitors than monomers in both cancer cell lines and in CMV.